The immobilization of test molecules on array supports has had a significant impact on drug discovery and medical diagnostic methods. The basic approaches for generating arrays of test molecules such as nucleic acid, protein or other organic molecules fall into two general categories. In the first such approach the test molecules are directly synthesized onto the array support, while in the second such approach the test molecules are attached to the support post-synthetically. Each approach has its own limitations and drawbacks. For example, when an array is created by direct synthesis onto an array support, the efficiency of each synthetic step affects the quality and integrity of molecules forming the array. The magnitude of the problem increases with the complexity of the individual molecules, potentially resulting in an undesirable percentage of incorrectly synthesized molecules and incomplete sequences. Such contaminants can interfere with subsequent use of the array. In contrast, the second approach to array production allows the desired molecules to be synthesized and purified by conventional methods prior to their formation into an array. Consequently, the quality of the arrayed molecules, and thus the quality of the resultant array, is potentially greater than that produced by the direct synthesis approach. However, a simple, effective patterning device for the preparation of arrays from pre-synthesized molecules is currently lacking.
The major challenges in constructing arrays of intermediate to high complexity (and/or density) include reagent/liquid confinement, volume control, position registration, the limited number of different array positions that can be simultaneously applied (parallelism), and speed. Several approaches to array production have been attempted with varying degrees of success.
Simple arrays have been constructed by manually pipetting a small amount of each desired test reagent onto a support, gradually building up an array. The limitations to such an approach are obvious. It is slow, few test reagent can be applied simultaneously, and position registration and spacing are poor. For example, Pastinen et al. (Genome Research, 7:606-614 (1997)) create an array of oligonucleotides by manually applying 0.5 .mu.L of a solution of 5'-amino-modified oligonucleotides onto an epoxide-activated glass slide to produce a 3.times.3 array of oligonucleotides on a 0.36 cm.sup.2 area of a preprinted glass slide.
Other, more traditional printing methods have been used to create patterns of a few different reagents on a solid support. Means such as silk screening, offset printing, and rotogravure printing have been used in the production of reagent test strips. In such methods, each reagent ink is applied separately. Thus, the number of different reagents that may be applied is limited. Furthermore, there is considerable potential for position registration difficulties. Johnson, for example, (U.S. Pat. No. 4,216,245) discusses methods for the production of reagent test strip devices.
Pipette dispensing of reagents can be automated. Automation potentially increases the speed and accuracy of array production, while decreasing the necessary spacing between array positions. However, the utility of automated pipetting methods are severely limited in the number of different reagents that may be simultaneously applied (low parallelism), and by the potential for aerosol formation from air bubbles. Since the array is built by the sequential addition of different reagents, producing a complex array is time consuming. Cozzette et al., for example, (U.S. Pat. No. 5,554,339) discusses the use of microsyringes for dispensing reagents during the production of biosensor devices.
High-speed robotics have also been used to print microarrays of amino-modified cDNA molecules onto silylated glass microscope slides (CEL Associates, Houston) or poly-1-lysine coated microscope slides (Schena, Bioassays, 18:427-431 (1996); Schena et al., Proc. Natl. Acad. Sci., U.S.A., 93:10614-10619 (1996).
Another approach to array printing is an adaptation of ink-jetting technology. For example, Hayes et al. (U.S. Pat. No. 4,877,745) discusses an ink-jet type method and apparatus for dispensing reagents, particularly in the production of reagent test strips.
Pin transfer is one approach that allows the simultaneous transfer of greater numbers of samples than possible with the above approaches
Pirrung et al., U.S. Pat. No. 5,143,854, Fodor et al., U.S. Pat. No. 5,510,270, Fodor et al., U.S. Pat. No. 5,445,934, and Chee et al., WO 95/11995) discuss the production of high density oligonucleotide arrays through a photolithographic synthesis of the oligonucleotides directly onto a derivatized glass substrate.
McGall et al. (U.S. Pat. No. 5,412,087) discusses a method for the production of a high density oligonucleotide array from pre-synthesized oligonucleotides.
Lebacq (U.S. Pat. No. 5,139,812) discusses the use of a pen, containing ink comprising a solution of a target nucleic acid, for cryptographically marking and thereby protecting valuable objects, such as art.
Despite all such methods, a method or apparatus capable of applying multiple "inks", containing different species of organic molecules, thereby creating an array, would be desirable. The present invention provides improved devices and methods for the production of arrays of molecules on solid or semi-solid supports. Previously, arrays could be constructed either manually, which is both time consuming and imprecise, or mechanically through the use of photolithographic, robotically-controlled or otherwise complex apparatus for the precise metering and placement of molecules. Alternatively, arrays could be constructed through direct chemical synthesis on a solid support. Such devices and methods have required undesirable tradeoffs between purity of the molecules forming the array, reagent confinement, volume control, position registration, parallelism, and speed. The present invention addresses the drawbacks of previous devices and methods, by providing a gene pen apparatus for the repetitive printing of arrays of molecules on solid or semi-solid supports.